Abstract Gamma-herpesvirus-driven lymphoproliferative disorders cause significant mortality in HIV+ populations. Despite highly active antiretroviral therapy, HIV+ individuals remain at elevated risk for Epstein-Barr virus (EBV)-associated B-cell cancers. EBV is identified in >40% of immunoblastic diffuse large B-cell lymphomas and nearly 100% of Hodgkin lymphoma and primary central nervous system lymphomas in HIV+ individuals. EBV transforms human B-cells into lymphoblastoid cell lines (LCL), a major model of EBV-driven immunoblastic lymphomas of immunosuppressed hosts. Yet, key EBV-induced host dependency factors remain to be elucidated. We therefore used systematic approaches to gain insights into key EBV oncoprotein targets. We used: 1) Genome-wide CRISPR screens to identify host dependency factors downstream of EBV oncoproteins; 2) Multiplexed mass spectrometry to create a proteomic map of EBV-mediated B-cell transformation; and 3) ChIP-seq to identify EBV oncoprotein host genomic targets, which found the first viral super-enhancers (SE), comprised of 5 LMP1- activated NF-kB and 4 EBV nuclear antigen subunits. These converged on the transcription factors BATF, IRF4 and IRF2 as key LMP1-targeted host dependency factors. Our preliminary studies indicate that BATF/IRF4 and IRF2 complexes have key dependency factor roles in EBV oncoprotein-driven MYC expression, necessary for LCL growth and survival. We propose to define how the interplay between viral oncoprotein superenhancers, BATF/IRF4 and IRF2 complexes drive MYC expression in lymphoblastoid B-cells. Our Specific Aims are to (1) Identify how EBV oncoproteins activate dependency factor BATF/IRF4 complexes to induce MYC; (2) Identify key BATF/IRF4 roles in EBV oncoprotein-induced MYC induction in lymphoblastoid B-cells; and (3) Identify how the viral SE-driven dependency factor IRF2 is necessary to support lymphoblastoid cell MYC expression. These studies specifically address PA-16-426 by advancing understanding of development, progression and treatment of malignancies observed in individuals with underlying HIV infection. The proposed studies are expected to contribute to understanding of how EBV oncoproteins and key genetically-defined downstream host dependency factors reprogram lymphoblastoid cell MYC expression. EBV oncoproteins may not be druggable, but SE and IRF4 are increasingly therapeutic targets. Since viral oncoprotein-induced MYC expression is critical for EBV-transformed B-cell growth, the longterm goal of these studies is to support rational approaches to restrain viral oncogene subversion of MYC.